The mechanism by which Mycoplasma pneumoniae produces Primary Atypical Pneumonia in man is not defined. Recent data indicate that the toxic factor is located in the cell membrane. The proposed investigation will examine the physiological and biochemical parameters in this membrane-associated cytopathic process. Hamster trachea organ cultures will be developed as the model system, with emphasis on the role of receptor sites and the kinetics of cell attachment. M. pneumoniae will be passaged repeatedly in vivo to obtain a highly virulent strain, and cell membranes will be fractioned to yield portions rich in "attachment tips" to see if they are responsible for virulence. Membranes will also be dissolved in detergents to separate lipid and protein, and then dialyzed to form reaggregated membranes. These will be assayed for cytotoxicity, as will hybrid membranes made from non- pathogenic Acholeplasma and pathogenic mycoplasma. When the relative importance of lipid and protein is known, membranes will be fractionated using column chromatography and polyacrylamide gel electrophoresis. This will be followed by the biochemical characterization of the toxin. This study of the membrane-associated toxin of M. pneumoniae and its interaction with epithelial cells will allow us to define the biochemical basis for virulence, and will thus contribute to our understanding of host-parasite relationships.